Complex systemic immune responses in humanized mice

The Jackson Laboratory presents its versatile toolkit of mouse models capable of supporting many functional components of the human immune system

Although many mammals have immune systems that share important characteristics with humans, certain immune cell types and phenomena are unique to us. As a result, it has been difficult to design and perform empirical studies that will help researchers accurately predict clinical responses to therapeutic drugs, infectious diseases, cancers, and other stimuli that can trigger an immune response. In particular, system-wide immune disorders such as cytokine release syndrome (CRS) and downstream graft-versus-host disease (GvHD) have remained incredibly difficult to study, prevent, and treat. In vitro studies do not capture the cascading effects that the immune system can have on a wide range of tissues, whereas live studies in animals such as rodents and non-human primates have never been successful in predicting toxicities related to single types of human immune cells (Eastwood et al., 2010).

To address these issues, the Jackson Laboratory (JAX) has developed a versatile set of mouse models capable of supporting many functional components of the human immune system. These mice provide a preclinical research platform that yields sensitive and reproducible results to screen drug candidates for inflammatory responses. The platform also offers insight into the progression of immune disorders and the potential effectiveness of drugs to treat them.

Humanize mice for immunological studies

To create our humanized mouse platform for studying T cell-based immune responses, we grafted immunodeficient mice (NOD-scid IL2rgnull, or NSG) with peripheral blood mononuclear cells (PBMC) from human donors. PBMCs include a variety of immune cells, but this engraftment model is dominated by T cells, allowing us to recreate a human T cell-mediated immune response in mice.

In validation studies, we found that we could reliably induce a dose-dependent response to CRS in these humanized mouse models using compounds known to clinically induce CRS, and that there was no CRS response to immunotherapies known to be clinically safe. Our results also captured the variation in cytokine release seen in individual human PBMC donors, with greater sensitivity and more reproducible results than in vitro tests performed with cells from the same donors (Ye et al., 2020).

Examine how CRS affects the organs

CRS occurs when the immune system overreacts to a stimulus such as a drug or disease. Activated white blood cells flood the body with excess cytokine signaling proteins, which activate more white blood cells and create a dangerous snowball effect that can lead to fever, organ failure, and even death.

With our humanized mouse model, we were able to not only measure the levels of various human cytokines released into the bloodstream following drug administration, but also observe the downstream effects of these cytokines on the body over time. We observed changes in body weight and collected serum to analyze kidney and liver enzyme levels. We also collected liver and lung tissue samples for histopathology studies using hematoxylin and eosin (H&E) staining, caspase 3 staining, and detection of single cell necrosis in the hepatocytes.


Figure 1. Comparison of the clinical serum half-life of three therapeutic antibodies with the corresponding half-life in a) WT mice or b) humanized FcRn (Tg32) mice, respectively. The R-squared of the linear regression model is shown in the graph, indicating a strong correlation between human PK of these molecules and that in humanized FcRn mice but not in the WT animal.

As expected, based on cytokine release levels, humanized mice treated with compounds known to cause CRS showed reduced body mass and suffered severe organ damage that reduced their functionality. We can continue to refine these studies to provide a better understanding of how factors such as variation in dosage can alter the effects of CRS on downstream organs.

Modeling the progression of GvHD

We also created humanized mice to model GvHD, which affects entire organ systems and can develop from CRS that has gone untreated for too long. We generated these models by treating NSG mice with irradiation and then engrafting them with human PBMCs, a process similar to our other studies. However, it was important that the PBMC donors for this particular study had already been pre-characterized specifically for their GvHD response.

We observed that the progression and severity of GvHD in humanized mouse models is specific to individual PBMC donors and closely reflects the diversity and range of severity of GvHD responses seen in human recipients. This indicates that, as with CRS, this platform could one day be used to help make individualized predictions about how patients’ immune systems will respond to treatment.

We carried out a first study to demonstrate the usefulness of this platform for the preclinical testing of new drugs intended to treat the complications of immune transplantation such as GvHD. We used the model to compare abatacept, an immunomodulator known to slow the progression of GvHD, to high and low doses of a bispecific antibody designed to accomplish the same thing.

Study complex systemic immune responses in humanized mice


Figure 1. Comparison of the clinical serum half-life of three therapeutic antibodies with the corresponding half-life in a) WT mice or b) humanized FcRn (Tg32) mice, respectively. The R-squared of the linear regression model is shown in the graph, indicating a strong correlation between human PK of these molecules and that in humanized FcRn mice but not in the WT animal.

In humans with GvHD, abatacept decreases T cell expansion and cytokine release and improves survival. Compared to controls, abatacept and bispecific antibody doses demonstrated all three of these effects. The high dose of the bispecific antibody performed better than abatacept in the final weeks of the study; this result demonstrates that this humanized mouse platform can provide nuanced insights into the efficacy and mode of action of any novel drug intended to treat GvHD.

References

Eastwood D, Findlay L, Poole S, Bird C, Wadhwa M, Moore M, Burns C, Thorpe R, Stebbings R. Monoclonal antibody TGN1412 trial failure explained by species differences in expression of CD28 on CD4+ effector memory T cells. Br J Pharmacol. October 2010;161(3):512-26. doi: 10.1111/j.1476-5381.2010.00922.x. PMID: 20880392

Ye C, Yang H, Cheng M, Shultz L, Greiner D, Brehm M, Keck J. A rapid, sensitive, and reproducible humanized PBMC in vivo mouse model for determining therapeutics-related cytokine release syndrome. FASB J. August 09, 2020;34:12963-12975. doi: 10.1096/fj.202001203R

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